Variant of nitrous oxide reductase protein and method of reducing concentration of nitrous oxide in sample

Abstract

Provided are polynucleotides encoding variants of nitrous oxide reductase proteins, recombinant microorganisms including a foreign gene encoding a variant of a nitrous oxide reductase protein, and a composition comprising the recombinant microorganism or the variant of a nitrous oxide reductase protein for use in removing nitrous oxide in a sample.

What is claimed is: 
     
         1 . A recombinant microorganism comprising a foreign gene encoding a variant of a nitrous oxide reductase protein, wherein the variant comprises an amino acid alteration of an amino acid residue corresponding to position L452 of the amino acid sequence of SEQ ID NO: 1 and has activity of an enzyme belonging to EC 1.7.2.4. 
     
     
         2 . The microorganism of  claim 1 , wherein the amino acid alteration is a substitution of the amino acid residue corresponding to position L452 with A, C, F, H, K, M, Q, or S. 
     
     
         3 . The microorganism of  claim 1 , wherein the amino acid alteration is an L452A, L452C, L452F, L452H, L452K, L452M, L452Q, or L4525 substitution. 
     
     
         4 . The microorganism of  claim 1 , wherein the variant further comprises an amino acid alteration of an amino acid residue corresponding to position H78 of the amino acid sequence of SEQ ID NO: 1. 
     
     
         5 . The microorganism of  claim 1 , further comprising a genetic modification that increases expression of a nosR gene encoding NosR, a nosD gene encoding NosD, a nosF gene encoding NosF, a nosY gene encoding NosY, and an apbE gene encoding ApbE. 
     
     
         6 . The microorganism of  claim 1 , wherein the microorganism belongs to the genus  Pseudomonas , the genus  Paracoccus , or the genus  Escherichia , or a combination thereof. 
     
     
         7 . A composition for reducing a concentration of nitrous oxide in a sample, the composition comprising a recombinant microorganism comprising a foreign gene encoding a variant of a nitrous oxide reductase protein, wherein the variant has an amino acid alteration of an amino acid residue corresponding to position L452 of the amino acid sequence of SEQ ID NO: 1 and has activity of an enzyme belonging to EC 1.7.2.4. 
     
     
         8 . The composition of  claim 7 , wherein the amino acid alteration is a substitution of the amino acid residue corresponding to position L452 with A, C, F, H, K, M, Q, or S. 
     
     
         9 . The composition of  claim 7 , wherein the amino acid alteration is an L452A, L452C, L452F, L452H, L452K, L452M, L452Q, or L4525 substitution. 
     
     
         10 . The composition of  claim 7 , wherein the variant further comprises an amino acid alteration of an amino acid residue corresponding to position H78 of the amino acid sequence of SEQ ID NO: 1. 
     
     
         11 . The composition of  claim 7 , wherein the microorganism further comprises a genetic modification that increases expression of a nosR gene encoding NosR, a nosD gene encoding NosD, a nosY gene encoding NosF, a nosY gene encoding NosY, and an apbE gene encoding ApbE. 
     
     
         12 . The composition of  claim 7 , wherein the microorganism belongs to the genus  Pseudomonas , the genus  Paracoccus , or the genus  Escherichia , or a combination thereof. 
     
     
         13 . A method of reducing a concentration of nitrous oxide in a sample, the method comprising contacting a sample comprising nitrous oxide, with a recombinant microorganism comprising a foreign gene encoding a variant of a nitrous oxide reductase protein or with a variant of a nitrous oxide reductase protein, wherein the variant comprises an amino acid alteration of an amino acid residue corresponding to position L452 of the amino acid sequence of SEQ ID NO: 1 and has activity of an enzyme belonging to EC 1.7.2.4. 
     
     
         14 . The method of  claim 13 , wherein the amino acid alteration is a substitution of the amino acid residue corresponding to position L452 with A, C, F, H, K, M, Q, or S. 
     
     
         15 . The method of  claim 13 , wherein the variant further comprises an amino acid alteration of an amino acid residue corresponding to position H78 of the amino acid sequence of SEQ ID NO: 1. 
     
     
         16 . The method of  claim 13 , wherein the microorganism further comprises a genetic modification that increases expression of a nosR gene encoding NosR, a nosD gene encoding NosD, a nosY gene encoding NosF, a nosY gene encoding NosY, and an apbE gene encoding ApbE. 
     
     
         17 . The method of  claim 13 , wherein the microorganism belongs to the genus  Pseudomonas , the genus  Paracoccus , or the genus  Escherichia , or a combination thereof. 
     
     
         18 . The method of  claim 13 , wherein the contacting comprises culturing or incubating the recombinant microorganism while contacting the recombinant microorganism with the sample comprising nitrous oxide. 
     
     
         19 . A variant of nitrous oxide reductase, comprising an amino acid alteration of an amino acid residue corresponding to position L452 of the amino acid sequence of SEQ ID NO: 1, the variant having activity of an enzyme belonging to EC 1.7.2.4, wherein the amino acid alteration is a substitution of the amino acid residue corresponding to position L452 with A, C, F, H, K, M, Q, or S. 
     
     
         20 . A polynucleotide encoding a variant of nitrous oxide reductase, wherein the variant comprises an amino acid alteration of an amino acid residue corresponding to position L452 of the amino acid sequence of SEQ ID NO: 1 and has activity of an enzyme belonging to EC 1.7.2.4, and wherein the amino acid alteration is a substitution of the amino acid residue corresponding to position L452 with A, C, F, H, K, M, Q, or S.